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1、HYBRIDOMA AND HYBRIDOMICS Volume 23, Number 2, 2004 Mary Ann Liebert, Inc. Murine Monoclonal Antibody to the SARS Coronavirus (SARS CoV) Spike Protein 137 ANTIGEN USED FOR IMMUNIZATION We used inactivated whole SARS CoV (coronavirus) as anti- gen (courtesy of Dr. Anton Andonov, National Microbiology
2、 Laboratory, Winnipeg, Canada). METHOD OF IMMUNIZATION For immunizations, 56-week-old female BALB/C mice were injected subcutaneously with 50 mg of inactivated SARS-CoV with an equal part of Complete Freunds Adjuvant (CFA, H37 Ra; Difco), on day 1. On day 30 the mice received 50 mg of purified SARS-
3、CoV in Incomplete Freunds Adjuvant (IFA). On days 48 and 63, mice received 5 mg in IFA. The mice re- ceived a final booster injection with 5 mg of purified SARS virus 3 days prior to fusion. Mice were euthanised by anaes- thesia overdose and exsanguinated by cardiac puncture. The spleens were excise
4、d under aseptic conditions. PARENTAL CELL LINE USED FOR FUSION P3X63 Ag8.653 SELECTION AND CLONING OF HYBRIDOMAS Immunization of mice, removal of spleens, preparation of spleen and myeloma cells, fusion and screening for monoclonal antibody to SARS were performed according to NCFAD stan- dard operat
5、ing procedures under ISO17025. Hybridomas were cloned out in semisolid medium. Supernatants were screened via ELISA using purified virus as antigen. Isotyping was per- formed using a commercial dipstick test (Roche) according to the manufacturers instructions. CloneClass F26G1IgG2a/k F26G6IgG2b/k F2
6、6G8IgG2a/k F26G18aIgG2b/k F26G19aIgG2a/k SPECIFICITY SARS-CoV spike protein SOURCE Murine FORMAT/PURITY Supernatants IMMUNOGEN Whole inactivated TOR-3 SARS CoV ANTIGEN REACTIVITY EIA, Western, IFA, IHC APPLICATIONS EIA, IHC, Western, IFA, cELISA, VNa METHOD OF IMMUNIZATION s.q. Monoclonal Antibodies
7、 STORAGE Frozen, 220 MEDIA BD Cell Quantum Yield AVAILABILITY SupernatantYes 3No ADDRESS CORRESPONDENCE TO: Dr. Jody D. Berry, Xin Yuan, Brigitte Nicolas, Fran Ranada, and Frank Plummer The Monoclonal Antibody Section National Centre for Foreign Animal Disease-CFIA National Microbiology LaboratoryHealth Canada Reagent Development Unit 1015 Arlington St. Winnipeg, MB, Canada Telephone: 204-789-6063 Fax: 204-789-2038 E-mail: berryjdinspection.gc.ca MONOCLONAL ANTIBODIES138
